Koontz Deborah, Baecher Kirsten, Kobrynski Lisa, Nikolova Stanimila, Gallagher Margaret
Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, Georgia.
Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, Georgia.
J Mol Diagn. 2014 Sep;16(5):533-540. doi: 10.1016/j.jmoldx.2014.05.003. Epub 2014 Jun 26.
The 22q11.2 deletion syndrome is one of the most common deletion syndromes in newborns. Some affected newborns may be diagnosed shortly after birth because of the presence of heart defects, palatal defects, or severe immune deficiencies. However, diagnosis is often delayed in patients presenting with other associated conditions that would benefit from early recognition and treatment, such as speech delays, learning difficulties, and schizophrenia. Fluorescence in situ hybridization (FISH) is the gold standard for deletion detection, but it is costly and time consuming and requires a whole blood specimen. Our goal was to develop a suitable assay for population-based screening of easily collectible specimens, such as buccal swabs and dried blood spots (DBS). We designed a pyrosequencing assay and validated it using DNA from FISH-confirmed 22q11 deletion syndrome patients and normal controls. We tested DBS from nine patients and paired buccal cell and venous blood specimens from 20 patients. Results were 100% concordant with FISH assay results. DNA samples from normal controls (n = 180 cell lines, n = 15 DBS, and n = 88 buccal specimens) were negative for the deletion. Limiting dilution experiments demonstrated that accurate results could be obtained from as little as 1 ng of DNA. This method represents a reliable and low-cost alternative for detection of the common 22q11.2 microdeletions and can be adapted to high-throughput population screening.
22q11.2缺失综合征是新生儿中最常见的缺失综合征之一。一些受影响的新生儿可能因存在心脏缺陷、腭裂或严重免疫缺陷而在出生后不久被诊断出来。然而，对于出现其他相关病症（如语言发育迟缓、学习困难和精神分裂症）的患者，诊断往往会延迟，而这些病症若能早期识别和治疗会有益处。荧光原位杂交（FISH）是检测缺失的金标准，但它成本高、耗时且需要全血标本。我们的目标是开发一种适用于基于人群筛查的检测方法，用于检测易于采集的标本，如口腔拭子和干血斑（DBS）。我们设计了一种焦磷酸测序检测方法，并使用来自FISH确诊的22q11缺失综合征患者和正常对照的DNA进行了验证。我们检测了9例患者的DBS以及20例患者的配对口腔细胞和静脉血标本。结果与FISH检测结果100%一致。正常对照的DNA样本（180个细胞系、15个DBS和88个口腔标本）缺失检测均为阴性。有限稀释实验表明，仅1 ng DNA就能获得准确结果。该方法是检测常见的22q11.2微缺失的一种可靠且低成本的替代方法，可适用于高通量人群筛查。
